• Journal of Life Science
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• v.26 no.8
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• pp.887-894
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• 2016

The extract from Artemisia annuain L.(AAE) is known as a medicinal herb that is effective against cancer. Apoptosis is the process of programmed cell death, and mitochondria are known to play a central role in cell death control. In this study, we evaluated the p53-independent apoptosis of extract of AAE through downregulation of Bcl-2 and the mitochondrial pathway in A549 (lung cancer cells). AAE may exert cancer cell apoptosis through regulating p-Akt, Cox-2, p53 and mitochondria-mediated apoptotic proteins. p-Akt/cox-2 is known to play an important role in cell proliferation and cell survival. The Bcl-2 pro-apoptotic proteins (such as Bax, Bak and Bim) mediate the permeabilization of the mitochondrial outer membrane. Treatment of AAE reduces p-Akt, p-Mdm2, cox-2 and anti-apoptotic proteins (such as Bcl-2), while tumor suppressor p53 and pro-apoptotic proteins. Activation of Bax/Bak releases cytochrome c from mitochondria to the cytosol to activate a caspase. Caspase-3 is the major effector caspase associated with apoptotic pathways. Caspase-3 generally exists in cytoplasm in the form of a pro-enzyme. In the initiation stage of apoptosis, caspase-3 is activated by proteolytic cleavage and activated caspase-3 cleaves poly (ADP-ribose) polymerase (PARP). We treated Pifithrin-α (p53 inhibitor) and Celecoxib (Cox-2 inhibitor) to learn the relationship between the signal transduction of proteins associated with apoptosis. These results suggest that AAE induces apoptosis through a p53-independent pathway in A549.

• Korean Journal of Medicinal Crop Science
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• v.17 no.2
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• pp.145-150
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• 2009

This study investigated the protective roles and mechanism of magnolol, from the stem bark of Magnolia officinalis against potential neurotoxin 3-hydroxykynurenine (3-HK)-induced neuronal cell death. For the evaluation of protective role of magnolol, we examined cell viability, apoptotic nuclei, change of mitochondrial membrane potential and caspase activity in human neuroblastoma SH-SY5Y cells. It was found that 3-HK induces neuronal cell death in the human neuroblastoma SH-SY5Y cell line. The reduced cell viability produced characteristic features such as cell shrinkages, plasma membrane blebbing, chromatin condensation, and nuclear fragmentation. The cells treated with 3-HK showed an increase in the concentration of reactive oxygen species (ROS) as well as in caspase activity. In addition, both are involved in the 3-HK-induced apoptosis. Magnolol attenuated the cell viability reduction by 3-HK in both a dose- and time-dependent manner. Optical microscopy showed that magnolol inhibited the cell morphological features in the 3-HK-treated cells. Furthermore, the increase in the ROS concentration and the caspase activities by 3-HK were also attenuated by magnolol. These results showed that magnolol has a protective effect on the 3-HK induced cell death by inhibiting ROS production and caspase activity.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.5
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• pp.1833-1837
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• 2015

An aristolactam-type alkaloid, isolated from Orophea enterocarpa, is enterocarpam-III (10-amino-2,3,4,6-tetramethoxyphenanthrene-1-carboxylic acid lactam). It is cytotoxic to various human and murine cancer cell lines; however, the molecular mechanisms remain unclear. The aims of this study were to investigate cytotoxic effects on and mechanism (s) of human cancer cell death in human hepatocellular carcinoma HepG2 and human invasive breast cancer MDA-MB-231 cells compared to normal murine fibroblast NIH3T3 cells. Cell viability was determined by MTT assay to determine $IC_{10}$, $IC_{20}$ and $IC_{50}$ levels, reactive oxygen species (ROS) production with 2',7'-dichlorohydrofluorescein diacetate and the caspase-3, -8 and -9 activities using specific chromogenic (p-nitroaniline) tetrapeptide substrates, viz., DEVD-NA, IETD-NA and LEHD-NA and employing a microplate reader. Mitochondrial transmembrane potential (MTP) was measured by staining with 3, 3'-dihexyloxacarbocyanine iodide ($DiOC_6$) and using flow cytometry. The compound was cytotoxic to HepG2 and MDA-MB-231 cells with the $IC_{50}$ levels of $26.0{\pm}4.45$ and $51.3{\pm}2.05{\mu}M$, respectively. For murine normal fibroblast NIH3T3 cells, the $IC_{50}$ concentration was $81.3{\pm}10.1{\mu}M$. ROS production was reduced in a dose-response manner in HepG2 cells. The caspase-9 and -3 activities increased in a concentration-dependent manner, whereas caspase-8 activity did not alter, indicating the intrinsic pathway activation. Enterocarpam-III decreased the mitochondrial transmembrane potential (MTP) dose-dependently in HepG2 cells, suggesting that the compound induced HepG2 cell apoptosis via the mitochondrial pathway. In conclusion, enterocarpam-III inhibited HepG2 and MDA-MB-231 cell proliferation and induced human HepG2 cells to undergo apoptosis via the intrinsic (mitochondrial) pathway and induction of caspase-9 activity.

• Nutrition Research and Practice
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• v.3 no.3
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• pp.180-184
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• 2009

The apoptotic effect of bacteria-derived $\beta$-glucan was investigated in human colon cancer cells SNU-C4 using terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay, reverse transcription-polymerase chain reaction (RT-PCR) expressions of Bcl-2, Bax, and Caspase-3 genes, and assay of caspase-3 enzyme activity. $\beta$-Glucan of 10, 50, and $100{\mu}g$/mL decreased cell viability in a dose-dependent manner with typical apoptotic characteristics, such as morphological changes of chromatin condensation and apoptotic body formation from TUNEL assay. In addition, $\beta$-glucan ($100{\mu}g$/mL) decreased the expression of Bc1-2 by 0.6 times, whereas the expression of Bax and Caspase-3 were increased by 3.1 and 2.3 times, respectively, compared to untreated control group. Furthermore, the caspase-3 activity in the $\beta$-glucan-treated group was significantly increased compared to those in control group (P < 0.05). Bacterial derived $\beta$-glucan could be used as an effective compound inducing apoptosis in human colon cancer.

• Journal of Physiology & Pathology in Korean Medicine
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• v.19 no.1
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• pp.212-217
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• 2005

The purpose of this study was to examine the effects of Citrus Reticulata(CR) on the Cell Detachment and Apoptosis in Human Gastric Cancer SNU-668 Cells. The effect of CR on apoptosis was investigated through MTT assay, DAPI staining, and TUNEL assay. We also performed RT-PCR for apoptotic genes including BCL-2, BAX, and caspase-3, the caspase-3 activity assay, and western blotting for pro-CASP-3. Then, to detect that adhesion of cell to ECM was reduced by CR, we investigated mRNA expression of CDH1 and PTK2 using RT-PCR, and their protein expressions using western blotting, and immunocytochemistry in SNU-668 cells. In this study, the results showed that treatment of CR induced time and dose-dependent cell death in SNU-668 cells. Downregulated mRNA expression of BCL-2, and upregulated mRNA expressions of BAX and CASP-3 indicated that the cell death was due to apoptosis. Protein expression of inactivated CASP-3, and caspase-3 activity assay also showed that apoptosis was induced in CR-treated cells.

• The Korean Journal of Physiology and Pharmacology
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• v.7 no.4
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• pp.199-205
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• 2003

Osteoblasts are affected by TNF-${\alpha}$ overproduction by immune cells during inflammation. It has been suggested that functional $NF-{\kappa}B$ sites are involved in TNF-${\alpha}$-induced bone resorption. Thus, we explored the effect of pyrrolidine dithiocarbamate (PDTC), which potently blocks the activation of nuclear factor $(NF-{\kappa}B)$, on the induction of TNF-${\alpha}$-induced activation of JNK/SAPK, AP-1, cytochrome c, caspase and apoptosis in MC3T3E1 osteoblasts. Pretreatment of the cells with PDTC blocked TNF-${\alpha}$-induced $NF-{\kappa}B$ activation. TNF-${\alpha}$-induced activation of AP-1, another nuclear transcription factor, was suppressed by PDTC. The activation of c-Jun N-terminal kinase, implicated in the regulation of AP-1, was also down regulated by PDTC. TNF-${\alpha}$-induced apoptosis, release of cytochrome c and subsequent activation of caspase-3 were abolished by PDTC. TNF-${\alpha}$-induced apoptosis was partially blocked by Ac-DEVD-CHO, a caspase-3 inhibitor, suggesting that caspase-3 is involved in TNF-${\alpha}$-mediated signaling through $NF-{\kappa}B$ in MC3T3E1 osteoblasts. Thus, these results demonstrate that PDTC, has an inhibitory effect on TNF-${\alpha}$-mediated activation of JNK/SAPK, AP-1, cytochrome c release and subsequent caspase-3, leading to the inhibition of apoptosis. Our study may contribute to the treatment of TNF-${\alpha}$-associated immune and inflammatory diseases such as rheumatoid arthritis and periodontal diseases.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.15
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• pp.6753-6759
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• 2015

Background: Loss of function of the p53 gene is implicated in defective apoptotic responses of tumors to chemotherapy. Although the pro-apoptotic roles of eugenol and capsaicin have been amply reported, their dependence on p53 for apoptosis induction in gastric cancer cells is not well elucidated. The aim of the study was to elucidate the role of p53 in the induction of apoptosis by eugenol and capsaicin in a human gastric cancer cell line, AGS. Materials and Methods: AGS cells were incubated with or without various concentrations of capsaicin and eugenol for 12 hrs, in the presence and absence of p53 siRNA. Cell cycling, annexin V and expression of apoptosis related proteins Bax, Bcl-2 ratio, p21, cyt c-caspase-9 association, caspase-3 and caspase-8 were studied. Results: In the presence of p53, capsaicin was a more potent pro-apoptotic agent than eugenol. However, silencing of p53 significantly abrogated apoptosis induced by capsaicin but not that by eugenol. Western blot analysis of pro-apoptotic markers revealed that as opposed to capsaicin, eugenol could induce caspase-8 and caspase-3 even in the absence of p53. Conclusions: Unlike capsaicin, eugenol could induce apoptosis both in presence and absence of functional p53. Agents which can induce apoptosis irrespective of the cellular p53 status have immense scope for development as potential anticancer agents.

• The Journal of Korean Obstetrics and Gynecology
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• v.31 no.2
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• pp.18-30
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• 2018

Objective: This study was designed to investigate the anti-cancer effects of AAH on ovarian cancer in vitro and by using allograft model in vivo. Methods: In this experiment, the effects of AAH on proliferation rates, cell morphology, cell death type, cell cycle, caspase activities and p38 mitogen-activated protein kinase (MAPK) pathway were investigated in A2780, human ovarian cell line. Results: AAH inhibited proliferation of A2780 cells in a dose dependent manner. In addition, AAH induced apoptosis but did not affect cell cycle of A2780 cells. AAH also effectively inhibited caspase 3 and caspase 9 activities respectively. In allograft tumor model, AAH reduced tumor volume and expanded life span in a dose dependent manner. Conclusion: It can be inferred that AAH can induce apoptosis in ovarian cancer cells and has possibility as an anticancer agent for ovarian cancer.

• Journal of the Korean Society of Food Science and Nutrition
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• v.35 no.5
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• pp.516-523
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• 2006

The mushroom Inonotus obliquue (IO) has been traditionally used for the treatment of gastrointestinal cancer in Russia, Poland, and most of Baltic countries. To explore the possibility that IO has chemoprevention effects, we examined whether or not the aqueous extract of IO inhibits HT-29 cell growth and investigated tile mechanism for this effect. Cells were incubated in the presence of increasing concentrations of the aqueous extract of IO. The extract substantially inhibited the viable HT-29 cell number in a dose-dependent manner and inhibited 5-bromo-2'-deoxyuridine incorporation into DNA of HT-29 cells. Annexin-V staining followed by flow cytometry revealed that the extract induced apoptosis of HT-29 cells in a dose-dependent manner. Western blot analysis of total cell lysates revealed that the extract induced cleavage of caspase-8, -9 and -3 and poly (ADP-ribose) polymerase, but did not affect the protein levels of Bax and Bcl-2. In addition, the extract dose-dependently increased the activity of caspase-8, -9 and -3. We have demonstrated that the aqueous extract of IO inhibits cell proliferation and induces apoptosis in HT-29 cells, which may be mediated by its ability to activate the caspase pathway.

• Journal of Integrative Natural Science
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• v.11 no.2
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• pp.93-100
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• 2018

Caspases, a family of cysteinyl aspartate-specific proteases plays a central role in the regulation and the execution of apoptotic cell death. Caspase-3 has been proven to be an effective target for reducing the amount of cellular and tissue damage because the activation of caspases-3 stimulates a signalling pathway that ultimately leads to the death of the cell. In this study, Hologram based Quantitative Structure Activity Relationship (HQSAR) models was generated on a series of Caspase-3 inhibitors named 3, 4-dihydropyrimidoindolones derivatives. The best HQSAR model was obtained using atoms, bonds, and hydrogen atoms (A/B/H) as fragment distinction parameter using hologram length 61 and 3 components with fragment size of minimum 5 and maximum 8. Significant cross-validated correlation coefficient ($q^2=0.684$) and non cross-validated correlation coefficients ($r^2=0.754$) were obtained. The model was then used to evaluate the eight external test compounds and its $r^2_{pred}$ was found to be 0.559. Contribution map show that presence of pyrrolidine sulfonamide ring and its bulkier substituent's makes big contributions for improving the biological activities of the compounds.